The invention relates to a method for purifying L-cysteine from an L-cysteine-containing fermentation broth.
L-Cysteine is an amino acid which, owing to the good solubility in water and the high sensitivity of the SH group to many reagents, e.g. to oxidants, can be purified and isolated only with great difficulty and with a high outlay.
U.S. Pat. No. 2,590,209 describes a general process for separating amino acid mixtures into acidic, neutral and basic amino acids, in which an acidic protein hydrolysate is firstly passed over a cation exchanger, the acidic and neutral amino acids are then eluted by means of ammonium hydroxide, while the basic amino acids are retained. These are subsequently flushed from the cation exchanger using a carbonate solution. The acidic and neutral amino acids are in turn separated by means of a downstream anion exchanger on which only the acidic amino acids are bound. The process described is not suitable for purifying cysteine since bases are used as eluent but cysteine is particularly oxidation-sensitive at high pH values.
Problems in the purification and also the prior art for isolation of L-cysteine are described in EP 1958933 A1 (corresponds to US 2008-0190854). This application describes a process in which an L-cysteine-containing fermentation broth comprising an oxidant which is able to oxidize L-cysteine at pH values of <5 is brought into contact at a pH of from 5 to 9 with an ion exchanger, where a pH of <5, preferably pH<2, occurs in the fermentation broth. Here, the L-cysteine binds to the ion exchanger and the bound L-cysteine is removed by means of an eluent from the ion exchanger. L-Cysteine monohydrochloride monohydrate can subsequently be obtained from the eluate by fractional crystallization. Chlorides of inorganic alkali metal and alkaline earth metal ions and also ammonium chloride are firstly crystallized by addition of concentrated HCl or by introduction of HCl gas. After filtration, the L-cysteine-containing mother liquor is cooled to −10° C. and L-cysteine hydrochloride monohydrate crystallizes. However, yield and purity are dependent on the purity of the L-cysteine-containing solution used; purities of up to >98% by weight require further fractional crystallization and/or recrystallization.
Processes for the fermentative preparation of L-cysteine are known, for example, from EP0885962B1 (corresponds to U.S. Pat. No. 5,972,663A). The processes allow inexpensive access to fermentation broths containing large amounts of L-cysteine.
Such an L-cysteine-containing fermentation broth is a complex mixture of substances. It contains not only L-cysteine but generally also L-cystine which is easily formed from L-cysteine under the conditions of fermentation, in particular by oxidation by means of oxygen present. In the presence of aldehydes or ketones, corresponding hemithioketals and/or thiazolidine derivatives of L-cysteine can also be present, as described, for example, in EP0885962B1. The fermentation broths can also contain small amounts of further amino acids or derivatives thereof. They generally also contain carbohydrates, salts of organic and inorganic cations and anions, e.g. alkali metal and alkaline earth metal salts and also traces of heavy metal salts (e.g. Fe, Cu, Mn, Zn, etc.), colorants and further impurities and additives, e.g. undesirable metabolites of the microorganisms used in the fermentation. Furthermore, the fermentation broths can also contain the raw materials and constituents used in the fermentation, e.g. customary C sources such as glucose, lactose, starch and the like, N sources such as ammonia/ammonium or proteins or protein hydrolysates and the like and also S sources such as sulfide, sulfite, sulfate, thiosulfate or dithionite and the like. Since L-cysteine is a sulfur-containing amino acid, an S source such as sulfide, sulfite, sulfate, thiosulfate or dithionite is generally fed in during the fermentation in order to provide a sufficient amount of the sulfur required for the formation of L-cysteine. Furthermore, dissolved oxygen is also present in the fermentation broths as a result of the oxygen introduced during the fermentation. The pH of these fermentation broths is usually 7, as described, for example, in EP0885962B1. Apart from the complexity of the composition of the fermentation broth, natural fluctuations in the ratio of the individual constituents additionally occur since the broth is a product of biological processes.